Antibody drug conjugates (ADC) are targeted chemotherapeutic molecules combining the properties of both antibodies and cytotoxic drugs by targeting potent cytotoxic drugs to antigen-expressing tumor cells, internalization, and release of drug, thereby enhancing their anti-tumor activity (Carter, P. and Senter, P. (2008) The Cancer Jour. 14(3):154-169).
Successful ADC development for a given target antigen depends on optimization of antibody selection, linker design and stability, cytotoxic drug potency and mode of drug and linker conjugation to the antibody (Dosio et al (2011) Toxins, 3:848-883; Polakis, P. (2005) Current Opinion in Pharmacology 5:382-387).
Certain pyrrolobenzodiazepine (PBD) compounds have the ability to recognize and bond to specific sequences of DNA; the preferred sequence is PuGPu (Pu=purine, such as adenine A and guanine G). The first PBD antitumor antibiotic, anthramycin, was discovered in 1965 (Leimgruber, et al., J. Am. Chem. Soc., 87:5793-5795 (1965); Leimgruber, et al., J. Am. Chem. Soc., 87, 5791-5793 (1965)). Since then, a number of naturally occurring PBD and analogues have been reported and described (Thurston, et al., Chem. Rev. 1994, 433-465 (1994); Antonow, D. and Thurston, D. E., (2011) Chem. Rev. 111 (4):2815-2864). Family members include abbeymycin (Hochlowski, et al., (1987) J. Antibiotics, 40:145-148), chicamycin (Konishi, et al., (1984) J. Antibiotics, 37:200-206), Thurston, et al., (1990) Chem. Brit., 26:767-772; Bose, et al., (1992) Tetrahedron, 48:751-758), mazethramycin (Kuminoto, et al., J. Antibiotics, 33, 665-667 (1980)), neothramycins A and B (Takeuchi, et al., J. Antibiotics, 29, 93-96 (1976)), porothramycin (Tsunakawa, et al., (1988) J. Antibiotics, 41:1366-1373), prothracarcin (Shimizu, et al, J. Antibiotics, (1982) 29:2492-2503; Langley and Thurston, (1987) J. Org. Chem., 52:91-97), sibanomicin, DC-102 (Hara, et al., (1988) J. Antibiotics, 41:702-704; Itoh, et al., J. Antibiotics, (1988) 41:1281-1284), sibiromycin (Leber, et al., (1988) J. Am. Chem. Soc., 110:2992-2993) and tomamycin (Arima, et al., (1972) J. Antibiotics, 25:437-444).
Pyrrolobenzodiazepines have the general structure:

and differ in the number, type and position of substituents, in both the aromatic A rings and pyrrolo C rings, and in the degree of saturation of the C ring. In the B-ring there is either an imine (N═C), a carbinolamine (NH—CH(OH)), or a carbinolamine methyl ether (NH—CH(OMe)) at the N10-C11 position, the electrophilic center responsible for alkylating DNA. All of the known natural products have an (9-configuration at the chiral C11a position which provides them with a right-handed twist when viewed from the C ring towards the A ring and determines the three-dimensional shape for isohelicity with the minor groove of B-form DNA, leading to a snug fit at the binding site (Kohn, In Antibiotics III. Springer-Verlag, New York, pp. 3-11 (1975); Hurley and Needham-VanDevanter, (1986) Acc. Chem. Res., 19:230-237). The ability of PBD to form an adduct in the minor groove, enables them to interfere with DNA processing, hence their use as antitumor agents. Pyrrolobenzodiazepine dimer compounds where two pyrrolobenzodiazepine structures are covalently attached by a linker through the C8 position of the A rings may dialkylate and crosslink double-stranded DNA (WO 2005/085251).
Pyrrolobenzodiazepine compounds can be employed as prodrugs by protecting them at the N10 position with a nitrogen protecting group, such as carbamate, which is removable in vivo (WO 2000/12507; WO 2005/023814). The protecting groups are removable from the N10 position of the PBD moiety to leave an N10-C11 imine bond. A range of protecting groups is described, including groups that can be cleaved by the action of enzymes.
Antibody-drug conjugates where the pyrrolobenzodiazepine (PBD) dimer is linked through the N10 position to an antibody specific for a tumor-associated antigen have in vitro and in vivo efficacy against tumor cells (WO 2011/130598). Antibody-drug conjugates with PBD dimer drug moieties having linker groups for connection to a cell binding agent, such as an antibody, via the bridge (“tether”) linking the monomer PBD units of the dimer have been described (WO 2007/085930). Antibody-drug conjugates with PBD dimer drug moieties having amide and amine groups in the B ring at N10-C11 position have been described (WO 2014/096368; WO 2013/177481; WO 2012/112708). Antibody drug conjugates comprising dialkylator pyrrolobenzodiazepine (PBD) dimer drug moieties linked at the N10 of the PBD by a disulfide linkage to antibodies have been described (WO2013/055987; Gregson et al. (2001) J. Med. Chem. 44:1161-1174).